Systematic Screening of Penetratin’s Protein Targets by Yeast Proteome Microarrays

Cell-penetrating peptides (CPPs) have distinct properties to translocate across cell envelope. The key property of CPPs to translocation with attached molecules has been utilized as vehicles for the delivery of several potential drug candidates that illustrate the significant effect in in-vitro experiment but fail in in-vivo experiment due to selectively permeable nature of cell envelop. Penetratin, a well-known CPP identified from the third α-helix of Antennapedia homeodomain of Drosophila, has been widely used and studied for the delivery of bioactive molecules to treat cancers, stroke, and infections caused by pathogenic organisms. Few studies have demonstrated that penetratin directly possesses antimicrobial activities against bacterial and fungal pathogens; however, the mechanism is unknown. In this study, we have utilized the power of high-throughput Saccharomyces cerevisiae proteome microarrays to screen all the potential protein targets of penetratin. Saccharomyces cerevisiae proteome microarrays assays of penetratin followed by statistical analysis depicted 123 Saccharomyces cerevisiae proteins as the protein targets of penetratin out of ~5800 Saccharomyces cerevisiae proteins. To understand the target patterns of penetratin, enrichment analyses were conducted using 123 protein targets. In biological process: ribonucleoprotein complex biogenesis, nucleic acid metabolic process, actin filament-based process, transcription, DNA-templated, and negative regulation of gene expression are a few significantly enriched terms. Cytoplasm, nucleus, and cell-organelles are enriched terms for cellular component. Protein-protein interactions network depicted ribonucleoprotein complex biogenesis, cortical cytoskeleton, and histone binding, which represent the major enriched terms for the 123 protein targets of penetratin. We also compared the protein targets of penetratin and intracellular protein targets of antifungal AMPs (Lfcin B, Histatin-5, and Sub-5). The comparison results showed few unique proteins between penetratin and AMPs. Nucleic acid metabolic process and cellular component disassembly were the common enrichment terms for penetratin and three AMPs. Penetratin shows unique enrichment items that are related to DNA biological process. Moreover, motif enrichment analysis depicted different enriched motifs in the protein targets of penetratin, LfcinB, Histatin-5, and Sub-5.

Melanoma Immunotherapy and Precision Medicine in the Era of Tumor Micro-Tissue Engineering: Where Are We Now and Where Are We Going?

Malignant melanoma still remains a cancer with very poor survival rates, although it is at the forefront of personalized medicine. Most patients show partial responses and disease progressed due to adaptative resistance mechanisms, preventing long-lasting clinical benefits to the current treatments. The response to therapies can be shaped by not only taking into account cancer cell heterogeneity and plasticity, but also by its structural context as well as the cellular component of the tumor microenvironment (TME). Here, we review the recent development in the field of immunotherapy and target-based therapy and how, in the era of tumor micro-tissue engineering, ex-vivo assays could help to enhance our melanoma biology knowledge in its complexity, translating it in the development of successful therapeutic strategies, as well as in the prediction of therapeutic benefits.

Diploid genome assembly of Kluyveromyces marxianus NRRL Y-50883 (SLP1)

The yeast Kluyveromyces marxianus SLP1 has the potential for application in biotechnological processes because it can metabolize several sugars and produce high-value metabolites. K. marxianus SLP1 is a thermotolerant yeast isolated from the mezcal process, and it is tolerant to several cell growth inhibitors such as saponins, furan aldehydes, weak acids, and phenolics compounds. The genomic differences between dairy and non-dairy strains related to K. marxianus variability are a focus of research attention, particularly the pathways leading this species toward polyploidy. We report the diploid genome assembly of K. marxianus SLP1 non-lactide strain into 32 contigs to reach a size of ∼12 Mb (N50 = 1.3Mb) and a ∼39% GC content. Genome size is consistent with the k-mer frequency results. Genome annotation by Funannotate estimated 5000 genes in haplotype A and 4910 in haplotype B. The enriched annotated genes by ontology show differences between alleles in biological processes and cellular component. The analysis of variants related to DMKU3 and between haplotypes shows changes in LAC12 and INU1, which we hypothesize can impact carbon source performance. This report presents the first polyploid K. marxianus strain recovered from non-lactic fermenting medium.

The Importance of the Extracellular Matrix in HPV-Associated Diseases

The extracellular matrix (ECM) is the non-cellular component of the tissues of our organism. It is the dynamic element that maintains a biochemical structure capable of supporting the organization and architecture of the tissue constituents. The diversity of ECM’s constituents gives it the biochemical and biophysical properties necessary to regulate its behavior and differentiation. ECM has an important role in the biology of cancer cell development and progression. Human papillomavirus infection (HPV) is the principal etiological agent of the most common sexually transmitted diseases. It is a virus that can cause lesions precursors of epithelial squamous and glandular tumors. Type 16 (HPV16) is the leading cause of pre-malignant lesions and invasive cancers in these tissues. This work will focus on HPV infection to understand the role of ECM in the invasion, spread, and pathogenesis of the lesions caused by this virus. Cancer is no longer considered a pathology explained only by uncontrolled proliferation and apoptosis but also by the deregulation of the microenvironment.

Targeting of MCL-1 in breast cancer associated fibroblasts reverses their myofibroblastic phenotype and pro-invasive properties

Cancer associated fibroblasts (CAF) are a major cellular component of epithelial tumors. In breast cancers in particular these stromal cells have numerous tumorigenic effects in part due to their acquisition of a myofibroblastic phenotype. Breast CAFs (bCAFS) typically express MCL-1. We show here that targeting this regulator of mitochondrial integrity using a specific BH-3 mimetic promotes fragmentation of these organelles without inducing cell death. MCL-1 antagonism in primary bCAFs directly derived from human samples mitigates myofibroblastic features and decreases expression of genes involved in actomyosin organization and contractility, associated with a cytoplasmic retention of the transcriptional regulator, Yes-Associated Protein (YAP). Such treatment decreases bCAFs ability to promote cancer cells invasion in 3D co-culture assays. These effects are counteracted by an inhibitor of the mitochondrial fission protein DRP-1, which interacts with MCL-1 upon BH3 mimetic treatment. Our findings underscore the usefulness of targeting MCL-1 in breast cancer ecosystems, not only to favor death of cancer cells but also to counteract the tumorigenic activation of fibroblasts with which they co-evolve.

Morphofunctional Rearrangement of the Fibrous Structures of the Rat Dermis under the Conditions of Implantation of 3D Scaffold Based on Polyprolactone

Background: the use of various scaffolds allows us to model the future fibrous framework of the newly formed regenerate, and also serves as a substrate for the settlement of the cellular component. The development of tissue engineering in regenerative medicine demands an understanding of the more specific mechanisms of the formation of the connective framework at the site of the defect. The aim of this research was to study the morphofunctional rearrangement of the fibrous structures of the rat dermis in response to the implantation of a 3D scaffold based on polyprolactone Methods and Results: The experiment was performed on 30 white male Wistar rats. The object of the study was a skin fragment together with an implantable 3D scaffold based on polyprolactone, taken on Days 3, 7 and 14 after implantation. Biomaterial with implantable scaffold was studied using light and scanning electron microscopy. The results of the study indicate that the 3D scaffold based on polyprolactone has good biocompatibility, causing a weak inflammatory reaction, and contributes to the formation of the connective tissue framework by Day 14. Conclusion: The results of the study can be used to develop new scaffolds or modify existing ones, as a "framework" for populating the cellular component and creating tissue-engineering structures.

Cadherin and Wnt signaling pathways as key regulators in diabetic nephropathy

Aim A recent meta-analysis of genome-wide linkage studies (GWLS) has identified multiple genetic regions suggestive of linkage with DN harboring hundreds of genes. Moving this number of genetic loci forward into biological insight is truly the next step. Here, we approach this challenge with a gene ontology (GO) analysis in order to yield biological and functional role to the genes, an over-representation test to find which GO terms are enriched in the gene list, pathway analysis, as well as protein network analysis. Method GO analysis was performed using protein analysis through evolutionary relationships (PANTHER) version 14.0 software and P-values less than 0.05 were considered statistically significant. GO analysis was followed by over-representation test for the identification of enriched terms. Statistical significance was calculated by Fisher’s exact test and adjusted using the false discovery rate (FDR) for correction of multiple tests. Cytoscape with the relevant plugins was used for the construction of the protein network and clustering analysis. Results The GO analysis assign multiple GO terms to the genes regarding the molecular function, the biological process and the cellular component, protein class and pathway analysis. The findings of the over-representation test highlight the contribution of cell adhesion regarding the biological process, integral components of plasma membrane regarding the cellular component, chemokines and cytokines with regard to protein class, while the pathway analysis emphasizes the contribution of Wnt and cadherin signaling pathways. Conclusions Our results suggest that a core feature of the pathogenesis of DN may be a disturbance in Wnt and cadherin signaling pathways, whereas the contribution of chemokines and cytokines need to be studied in additional studies.

Multi-Level Analyses of Genome-Wide Association Study to Reveal Significant Risk Genes and Pathways in Neuromyelitis Optica Spectrum Disorder

BackgroundNeuromyelitis optica spectrum disorder (NMOSD) is an inflammatory disease of the central nervous system and it is understandable that environmental and genetic factors underlie the etiology of NMOSD. However, the susceptibility genes and associated pathways of NMOSD patients who are AQP4-Ab positive and negative have not been elucidated.MethodsSecondary analysis from a NMOSD Genome-wide association study (GWAS) dataset originally published in 2018 (215 NMOSD cases and 1244 controls) was conducted to identify potential susceptibility genes and associated pathways in AQP4-positive and negative NMOSD patients, respectively (132 AQP4-positive and 83 AQP4-negative).ResultsIn AQP4-positive NMOSD cases, five shared risk genes were obtained at chromosome 6 in AQP4-positive NMOSD cases by using more stringent p-Values in both methods (p < 0.05/16,532), comprising CFB, EHMT2, HLA-DQA1, MSH5, and SLC44A4. Fifty potential susceptibility gene sets were determined and 12 significant KEGG pathways were identified. Sixty-seven biological process pathways, 32 cellular-component pathways, and 29 molecular-function pathways with a p-Value of <0.05 were obtained from the GO annotations of the 128 pathways identified. In the AQP4 negative NMOSD group, no significant genes were obtained by using more stringent p-Values in both methods (p < 0.05/16,485). The 22 potential susceptibility gene sets were determined. There were no shared potential susceptibility genes between the AQP4-positive and negative groups, furthermore, four significant KEGG pathways were also identified. Of the GO annotations of the 165 pathways identified, 99 biological process pathways, 37 cellular-component pathways, and 29 molecular-function pathways with a p-Value of <0.05 were obtained.ConclusionThe potential molecular mechanism underlying NMOSD may be related to proteins encoded by these novel genes in complements, antigen presentation, and immune regulation. The new results may represent an improved comprehension of the genetic and molecular mechanisms underlying NMOSD.